The long-range goal of this project is to dissect the hedgehog signaling pathway and determine its role in forebrain development. Our studies focus on the genetic and molecular basis for the formation of the ventral diencephalon and the associated anterior pituitary. We have identified four zebrafish mutations that affect the development of this embryonic region: sonic you, detour, you-too, and chameleon. The sonic you mutation disrupts the zebrafish sonic hedgehog gene, and our recent studies have identified detour and you-too as mutations in zebrafish gli1 and gli2, respectively. Members of the gli gene family encode transcription factors that are thought to mediate hedgehog signaling. The finding that three of the four diencephalic genes affect components of the hedgehog signaling pathway establishes the important role of this universal signaling cascade during forebrain development. We propose to further determine the function and regulatory interactions of the hedgehog signaling pathway during the development of ventral diencephalon and anterior pituitary by A) conducting a detailed analysis of diencephalic and pituitary development in wild-type and sonic you mutant embryos using marker gene expression and fate map analysis; B) studying the transcriptional and post-translational regulation of gli factor activity by hedgehog signaling; C) cloning the chameleon gene. Abnormal diencephalic and pituitary development has been implicated in human congenital disorders such as holoprosencephaly, and components of the hedgehog signaling pathway are involved in human cancer. The proposed studies will thus help to provide the necessary context for understanding birth defects and cancer formation in humans.